The goal of this proposal is to discover novel cancer-causing somatic mutations in lung squamous cell carcinoma (SCC) with a combination of multiple sequencing technologies and functional experiments. Lung SCC accounts for 30% of all lung cancer cases. Unlike other types of lung cancer, there are no targeted therapies available for lung SCC patients. It is hoped that an accurate characterization of the spectrum of mutations in Lung SCC will lead to new avenues of rational drug therapy in lung SCC. Aim 1: Determine frequency of newly discovered somatic mutations in lung SCC. In preliminary experiments, I analyzed DNA sequence data for 201 genes and 496 microRNAs in 20 lung SCC samples. This study discovered 27 somatic point mutations across 17 different genes. Examination of the mutations revealed several potentially interesting findings. In order to determine which genes are recurrently mutated in lung squamous cancer and thus likely to be involved in tumorigenesis, Sanger sequencing of select mutated genes will be performed in an expanded set of 96 primary lung SCC samples. Aim 2: Utilize single-template Solexa sequencing for discovery of fusion genes. Fusion genes often drive malignancy in many hematological cancers. Importantly, fusion genes such as BCR-ABL can be effectively treated with imatinib. Recent findings in other cancers suggest that fusion genes could also play a significant role in epithelial cancers. TO discover fusion genes in lung SCC, I will perform Solexa shotgun sequencing of cDNA in lung SCC cell line models. Aim 3: Validate mutation and fusion gene candidates from Aims 1 and 2 with in vitro transformation assays. In order to determine which alterations from Aims 1 and 2 are truly involved in pathogenesis, I will systematically test discovered mutations and fusion genes with transformation assays and RNAi experiments in in vitro cell line models. This proposal will attempt to discover mutations in lung cancer patients by analyzing DNA from tumor samples. It is hoped that newly found mutations will provide the stepping stone for future lung cancer therapies.